PCR detection of Angiostrongylus vasorum in faecal samples of dogs and foxes

Abstract

The cardiovascular nematode Angiostrongylus vasorum is spreading in the fox and dog populations of northern Europe. A. vasorum can result in severe clinical manifestations in dogs; therefore, specific diagnosis is crucial for assessing its prevalence. In the present study, faecal samples from foxes and domestic dogs were tested by a new polymerase chain reaction (PCR) targeting the second internal transcribed region of the ribosomal DNA (ITS2) of A. vasorum. Initial isolation of faecal larvae by sieving facilitated the processing of larger sample volumes and allowed for the recovery of dead larvae from frozen samples. The sieve-PCR method enabled the identification of a single larva per 2 g of faecal sample and did not amplify DNA of a range of canine helminths, thus presenting a non-invasive tool for wildlife surveillance and for confirmative diagnosis in dogs. Abstract The cardiovascular nematode Angiostrongylus vasorum is spreading in the fox and dog populations of northern Europe. A. vasorum can result in severe clinical manifestations in dogs; therefore, specific diagnosis is crucial for assessing its prevalence. In the present study, faecal samples from foxes and domestic dogs were tested by a new polymerase chain reaction (PCR) targeting the second internal transcribed region of the ribosomal DNA (ITS2) of A. vasorum. Initial isolation of faecal larvae by sieving facilitated the processing of larger sample volumes and allowed for the recovery of dead larvae from frozen samples. The sieve-PCR method enabled the identification of a single larva per 2 g of faecal sample and did not amplify DNA of a range of canine helminths, thus presenting a non-invasive tool for wildlife surveillance and for confirmative diagnosis in dogs.

Calodium hepaticum (Nematoda: Capillaridae) in a red fox (Vulpes vulpes) in Italy with scanning electron microscopy of the eggs

“…Morphological and morphometric analyses of eggs were performed with light and scanning electron microscopy (SEM). For SEM, eggs were isolated by flotation in a zinc sulphate solution (s.g. 1.350) and sieved, following the technique described in Al-sabi et al (2010), then mounted on aluminium stubs, air dried, gold coated with the sputtering technique, observed and photographed.…”

Serological Detection of Circulating Angiostrongylus vasorum Antigen- and Parasite-Specific Antibodies in Dogs from Poland

“…requires expertise (McGarry and Morgan 2009). Alternative diagnostic tools such as molecular methods Al-Sabi et al 2010) or highly specific serological tests have been developed (Schnyder et al 2011b;Schucan et al 2012) to overcome these problems. ELISAs represent a valid and affordable tool for diagnosis in both, individual and population studies, and have been very recently validated in a field study (Guardone et al 2013).…”

The first clinically manifested case of angiostrongylosis in a dog in Slovakia

“…PCR reaction for the amplification of rDNA second internal transcribed spacer (ITS2) was performed in a final volume of 25 μl using specific A. vasorum pairs of primers AV5 and AV4 designed by Al-Sabi et al (2010). Template DNA (5 µl) was amplified by PCR reaction started with a pre-heating step at 94°C for 2 min and 38 cycles consisted of denaturing at 94°C for 30 s, annealing at 57°C for 30 s and extension at 72°C for 30 s. The PCR was terminated with a final extension at 72°C for 7 min.…”